Tag Archives: smooth muscle

Calmodulin antagonists effect on Ca(2+) level in the mitochondria and cytoplasm of myometrium cells

S. G. Shlykov, L. G. Babich, M. E. Yevtushenko, S. O. Karakhim, S. O. Kosterin

Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: sshlykov@biochem.kiev.ua

It is known that Са2+-dependent regulation of this cation exchange in mitochondria is carried out with participation of calmodulin. We had shown in a previous work using two experimental models: isolated mitochondria and intact myometrium cells, that calmodulin antagonists reduce the level of mitochondrial membrane polarization. The aim of this work was to investigate the influence of calmodulin antagonists on the level of ionized Са in mitochondria and cytoplasm of uterine smooth muscle cells using spectrofluorometry and confocal microscopy. It was shown that myometrium mitochondria, in the presence of АТР and MgCl2 in the incubation medium, accumulate Са ions in the matrix. Incubation of mitochondria in the presence of СССР inhibited cation accumulation, but did not cease it. Calmodulin antagonist such as trifluoperazine (100 µМ) considerably increased the level of ionized Са in the mitochondrial matrix. Preliminary incubation of mitochondria with 100 µМ Са2+, before adding trifluoperazine to the incubation medium, partly prevented influence of the latter on the cation level in the matrix. Incubation of myometrium cells (primary culture) with another calmodulin antagonist calmidazolium (10 µМ) was accompanied by depolarization of mitochondrial membrane and an increase in the concentration of ionized Са in cytoplasm. Thus, using two models, namely, isolated mitochondria and intact myometrium cells, it has been shown that calmodulin antagonists cause depolarization of mitochondrial membranes and an increase of the ionized Са concentration in both the mitochondrial matrix and the cell cytoplasm.

Protective effect of tiacalix[4]arene-tetrasulphonate on heavy metal inhibition of myometrium myosin subfragment-1 ATP-hydrolase activity

R. D. Labyntsevа1, O. V. Bevza1, A. A. Bevza1, A. M. Lulko1,
S. Kharchenko2, V. I. Kalchenko2, S. O. Kosterin1

1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
е-mail: labyntseva@biochem.kiev.ua;  kinet@biochem.kiev.ua;
2Institute of Organic Chemistry, National Academy of Sciences of Ukraine, Kyiv;
е-mail: vik@ioch.kiev.ua

Heavy metals have a negative effect on the contractility of uterine smooth muscles (myometrium), these effects can lead to various pathologies of a women reproductive system. To overcome these effects the methods for correcting the myometrium contractile activity are  to be developed. Catalyzed by myosin ATPase ATP hydrolysis is the most important reaction in the molecular mechanism of myo­metrium contraction. We have found an inhibitory effect of 0.03-0.3 mM Ni2+, Pb2+ and Cd2+ on enzymatic hydrolysis of ATP by myosin subfragment-1 obtained from swine uterine smooth muscles. We have demonstrated that 100 µM thiacalix[4]arene-tetrasulphonate (C-798) recovered to the control level of ATPase activity of myosin subfragment-1 in the presence of heavy metal cations. One of the most probable mechanisms of C-798 corrective activity is based on its ability to chelate heavy metals, thus cations Pb, Cd and Ni can be removed from the incubation medium. Computer simulation has demonstrated that the protective effect of C-798 may also be the result of weakening the interaction of heavy metal ions with amino acid residues of the myosin molecule near the active site of ATP hydrolase. The obtained results can be used for further research aimed at assessing the prospects of thiacalix[4]arene-tetrasulfonate as pharmacological compounds.

Ca(2+)/H(+)-exchange in myometrium mitochondria

O. V. Kolomiets, Yu. V. Danylovych, H. V. Danylovych, S. O. Kosterin

Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: danylovych@biochem.kiev.ua

Using the fluorescent probe Fluo-4 AM the authors have identified Na+-independent Ca2+/H+-exchange in isolated mitochondria of rat myometrium and studied its individual properties.  Formation of directional protons gradient in the matrix of mitochondria causes antyporte release of Ca2+, which has been previously accumulated in energetic processes (in the presence of Mg-ATP and succinate). The functioning of Ca2+/H+-exchange depends on the proton gradient and is characterized by reversibility, in case of extramitochondria environment alkalization the additional accumulation of Ca2+ by organelles is recorded. Monovalent cations gradients (Na+, K+, Li+) do not cause the release of Ca2+ from mitochondria. Rate of Ca2+/H+-exchange is growing in terms of increasing ΔpH on the mitochondria membrane and kinetics of ΔpH-induced Ca2+ release from the matrix corresponds to the laws of first order reaction.  Research of Ca2+/H+-exchange some properties in the myometrium mitochondria showed that the above transport process is of electrogenic nature, perhaps it is done in a 1 : 1 stechiometry (Hill coefficient on H+ close to 1) and is able to adjust matrix Ca2+ concentration under physiological conditions (pH activation of about 6.9).  Thus, in the inner membrane of the myometrium mitochondria the available system of the seconda­ry active Ca2+-transport from the matrix of these organelles to myoplasm and the functioning of Ca2+/H+-exchanger may underlie this process.